Variant-specific methylation of H1 by G9a/Glp1. (a) Alignment of H1 variants H1.2, H1.3, H1.4, H1.5 and H1.0. Amino acids identical in at least three variants are highlighted in blue. The regions around H1.4K26 and H1.2K187 are boxed. (b) H1.4K26 and H1.2K187 are G9a and Glp1 targets. Methylation assay on indicated H1.2 and H1.4 mutants using (left) immunoprecipitated Glp1 and (right) G9a. Note that the high expression of H1 led to partial N-terminal degradation of some constructs in bacteria. (Upper panels) Ponceau (Pon) staining: *degradation fragments. (Lower panels) Autoradiography (Rad). (c) H1.2, H1.3, H1.4, H1.5 and H1.0 were methylated by G9a and Glp1. Methylation assays on recombinant human H1 variants using immunoprecipitated G9a and Glp1. (Upper panels) Ponceau (Pon) staining; (lower panels) autoradiography (Rad). (d) Methylation assays on human recombinant H1 followed by chymotrypsin digestion, which separates the N-terminus and core from the C-terminus. Only the N-terminus and core of H1.4 is a major G9a/Glp1 target whereas H1.2, H1.3, H1.5 and H1.0 are methylated by G9a/Glp1 preferentially in the C-terminal part. Autoradiography (Rad) is shown (loading control in Additional file 3).